Cross-talk between calcium-calmodulin and nitric oxide in abscisic acid signaling in leaves of maize plants

Using pharmacological and biochemical approaches, the signaling pathways between hydrogen peroxide （H2O2）, calcium （Ca^2＋）-calmodulin （CAM）, and nitric oxide （NO） in abscisic acid （ABA）-induced antioxidant defense were investigated in leaves of maize （Zea mays L.） plants. Treatments with ABA, H2O2, and CaCl2 induced increases in the generation of NO in maize mesophyll cells and the activity of nitric oxide synthase （NOS） in the cytosolic and microsomal fractions of maize leaves. However, such increases were blocked by the pretreatments with Ca^2＋ inhibitors and CaM antagonists. Meanwhile, pretreatments with two NOS inhibitors also suppressed the Ca^2＋-induced increase in the production of NO. On the other hand, treatments with ABA and the NO donor sodium nitroprusside （SNP） also led to increases in the concentration of cytosolic Ca^2＋ in protoplasts of mesophyll cells and in the expression of calmodulin 1 （CaM1） gene and the contents of CaM in leaves of maize plants, and the increases induced by ABA were reduced by the pretreatments with a NO scavenger and a NOS inhibitor. Moreover, SNP-induced increases in the expression of the antioxidant genes superoxide dismutase 4 （SOD4）, cytosolic ascorbate peroxidase （cAPX）, and glutathione reductase 1 （GR1） and the activities of the chloroplastic and cytosolic antioxidant enzymes were arrested by the pretreatments with Ca^2＋ inhibitors and CaM antagonists. Our results suggest that Ca^2＋-CaM functions both upstream and downstream of NO production, which is mainly from NOS, in ABA- and H2O2-induced antioxidant defense in leaves of maize plants.     

Involvement of Polyamine Oxidase in Abscisic Acidinduced Cytosolic Antioxidant Defense in Leaves of Maize

Using pharmacological and biochemical approaches, the role of maize polyamine oxidase （MPAO） in abscisic acid （ABA）- induced antioxidant defense in leaves of maize （Zea mays L.） plants was investigated. Exogenous ABA treatment enhanced the expression of the MPAO gene and the activities of apoplastic MPAO. Pretreatment with two different inhibitors for apoplastic MPAO partly reduced hydrogen peroxide （H202） accumulation induced by ABA and blocked the ABA-induced expression of the antioxidant genes superoxide dismutase 4 and cytosolic ascorbate peroxidase and the activities of the cytosolic antioxidant enzymes. Treatment with spermidine, the optimum substrate of MPAO, also induced the expression and the activities of the antioxidant enzymes, and the upregulation of the antioxidant enzymes was prevented by two inhibitors of MPAO and two scavengers of H202. These results suggest that MPAO contributes to ABA-induced cytosolic antioxidant defense through H202, a Spd catabolic product.     

Production of Polyamines Is Enhanced by Endogenous Abscisic Acid in Maize Seedlings Subjected to Salt Stress

It is known that salt stress and exogenously applied abscisic acid （ABA） can enhance the polyamine content in plants and that salt stress itself can lead to an increase in endogenous ABA production. In the present study, the relationships between salt-induced ABA and polyamine accumulation were inves- tigated using ABA-deficient mutant （vp5/vp5） maize （Zea mays L.） seedlings and ABA and polyamine biosynthesis inhibitors. The results show that reduced endogenous ABA levels, as a result of either the mutation or by using a chemical inhibitor （sodium tungstate）, also reduced the accumulation of polyamines in salt-stressed leaves of maize seedlings. The polyamine synthesis inhibitors D-arginine and α- difluoromethylornithine also reduced the polyamine content of the leaves of maize seedling under salt stress. Both ABA and polyamine enhanced the dry weight accumulation of salt-stressed seedlings and also increased the activities of the two dominant tonoplast membrane enzymes, H^＋-ATPase and H^＋-PPase, when plants were under salt stress. The results suggest that salt stress induces an increase in endogenous ABA levels, which then enhances polyamine synthesis. Such responses may increase a plant＇s tolerance to salt.     

Nitric oxide reduces hydrogen peroxide accumulation involved in water stress-induced subcellular anti-oxidant defense in maize plants

Nitric oxide （NO） is a bioactive molecule involved in many biological events, and has been reported as pro-oxidant as well as anti-oxidant in plants. In the present study, the sources of NO production under water stress, the role of NO in water stress-induced hydrogen peroxide （H2O2） accumulation and subcellular activities of anti-oxidant enzymes in leaves of maize （Zea mays L.） plants were investigated. Water stress induced defense increases in the generation of NO in maize mesphyll cells and the activity of nitric oxide synthase （NOS） in the cytosolic and microsomal fractions of maize leaves. Water stress-induced defense increases in the production of NO were blocked by pretreatments with inhibitors of NOS and nitrate reductase （NR）, suggesting that NO is produced from NOS and NR in leaves of maize plants exposed to water stress. Water stress also induced increases in the activities of the chloroplastic and cytosolic anti-oxidant enzymes superoxide dismutase （SOD）, ascorbate peroxidase （APX）, and glutathione reductase （GR）, and the increases in the activities of anti-oxidant enzymes were reduced by pretreatments with inhibitors of NOS and NR. Exogenous NO increases the activities of water stress-induced subcellular anti-oxidant enzymes, which decreases accumulation of H2O2. Our results suggest that NOS and NR are involved in water stress-induced NO production and NOS is the major source of NO. The potential ability of NO to scavenge H2O2 is, at least in part, due to the induction of a subcellular anti-oxidant defense.     

Carbon Monoxide Alleviates Salt-Induced Oxidative Damage in Wheat Seedling Leaves

Carbon monoxide （CO）, a by-product released during the degradation of heme by heme oxygenases （EC 1.14.99.3） In animals, is regarded as an important physiological messenger or bioactive molecule involved in many biological events that has been recently reported as playing a major role in mediating the cytoprotectlon against oxidant-induced lung Injury. In the present study, we first determined the protective effect of exogenous CO against salt-induced oxidative damage in wheat seedling leaves. Wheat seedlings treated with 0.01μmol/L hematin as the CO donor demonstrated significant reversal of chlorophyll decay, dry weight, and water loss induced by 300 mmol/L NaCl stress. Interestingly, the increase in lipid peroxidation observed in salt-treated leaves was reversed by 0.01μmol/L hematin treatment. Time-couree analyses showed that application of 0.01μmol/L hematln enhanced gualacol peroxidase, superoxide dismutase, ascorbate peroxidase and catalase activities in wheat seedling leaves subjected to salt stress. These effects are specific for CO because the CO scavenger hemoglobin （1.2 mg/L） blocked the actions of the CO donor hematln. However, higher concentration of the CO donor （1.0μmol/L） did not alleviate dry weight and water loss of salt-stressed wheat seedlings. These results suggest that exogenous application of low levels of a CO donor may be advantageous against salinity toxicity.     

Pyrrolidine dithiocarbamate protects pancreatic β-cells from oxidative damage through regulation of FoxO1 activity in type 2 diabetes rats

Pyrrolidine dithiocarbamate （PDTC） can lower the bloot glucose level and improve the insulin sensitivity in diabeti, rats. However, the mechanisms underlying this effect o PDTC treatment in diabetic rats remained uncertain, h this study, we evaluated the mechanisms by which PDT（ conferred protection against oxidative damage to pancreat ic islet β-cells in rats with experimental type 2 diabete mellitus （DM）. DM in the rats was elicited by long-tern high-fat diet accompanied with a single intraperitonea （i.p.） injection of a low dose of streptozotocin. After a 7-da1 administration of PDTC （50 mg/kg/day i.p.）, blood glucos levels were measured and pancreatic tissues were collecte / for the determination of various biochemical and enzyma 1 ic activities using immunohistochemistry, immunofluoresI cence, and western blot techniques. The percentage o 1 apoptotic pancreatic islet β-cells was detected by flow cyto metry. The results showed that diabetic rats had elevate blood glucose levels and insulin resistance, accompanieq with an increase in malondialdehyde content, nitrotyrosin production, and inducible nitric oxide synthase expression A decrease in superoxide dismutase and glutathione pero idase activities was also observed in DM rats, culminatin with elevated β-cell apoptosis. PDTC treatment significantl reduced the oxidative damage and the β-cell apoptosi and also increased the insulin production through down-reg lating FoxO1 acetylation and up-regulating nuclear PDX- level. These data suggested that PDTC can protect islet βcells from oxidative damage and improve insulin productio through regulation of PDX-1 and FoxO1 in a DM rat model.     

Evaluation of spatial and temporal patterns of insect damage and aflatoxin level in the pre-harvest corn fields to improve management tactics

Spatial and temporal patterns of insect damage in relation to aflatoxin contamination in a corn field with plants of uniform genetic background are not well understood. After previous examination of spatial patterns of insect damage and aflatoxin in pre-harvest corn fields, we further examined both spatial and temporal patterns of cob- and kernel- feeding insect damage, and aflatoxin level with two samplings at pre-harvest in 2008 and 2009. The feeding damage by each of the ear/kernel-feeding insects （i.e., corn earworm/fall armyworm damage on the silk/cob, and discoloration of corn kernels by stink bugs） and maize weevil population were assessed at each grid point with five ears. Sampling data showed a field edge effect in both insect damage and aflatoxin contamination in both years. Maize weevils tended toward an aggregated distribution more frequently than either corn earworm or stink bug damage in both years. The frequency of detecting aggregated distribution for aflatoxin level was less than any of the insect damage assessments. Stink bug damage and maize weevil number were more closely associated with aflatoxin level than was corn earworm damage. In addition, the indices of spatial-temporal association （χ） demonstrated that the number of maize weevils was associated between the first （4 weeks pre-harvest） and second （1 week pre-harvest） samplings in both years on all fields. In contrast, corn earworm damage between the first and second samplings from the field on the Belflower Farm, and aflatoxin level and corn earworm damage from the field on the Lang Farm were dissociated in 2009.     

Shoot-Specific Down-Regulation of Protein Farnesyltransferase （α-Subunit） for Yield Protection against Drought in Canola

Canola （Brassica napus L.） is one of the most important oilseed crops in the world and its seed yield and quality are significantly affected by drought stress. As an innate and adaptive response to water deficit, land plants avoid potential damage by rapid biosynthesis of the phytohormone abscisic acid （ABA）, which triggers stomatal closure to reduce transpirational water loss. The ABA-mediated stomatal response is a dosage-dependent process; thus, one genetic engineering approach for achieving drought avoidance could be to sensitize the guard cell＇s responsiveness to this hormone. Recent genetic studies have pinpointed protein farnesyltransferase as a key negative regulator controlling ABA sensitivity in the guard cells. We have previously shown that down-regulation of the gene encoding Arabidopsis β-subunit of farnesyltransferase （ERA1） enhances the plant＇s sensitivity to ABA and drought tolerance. Although the β-subunit of famesyltransferase （AtFTA） is also implicated in ABA sensing, the effectiveness of using such a gene target for improving drought tolerance in a crop plant has not been validated. Here, we report the identification and characterization of the promoter of Arabidopsis hydroxypyruvate reductase （AtHPR1）, which expresses specifically in the shoot and not in non-photosynthetic tissues such as root. The promoter region of AtHPR1 contains the core motif of the well characterized dehydration-responsive cis-acting element and we have confirmed thatAtHPR1 expression is inducible by drought stress. Conditional and specific down-regulation of FTA in canola using the AtHPR1 promoter driving an RNAi construct resulted in yield protection against drought stress in the field. Using this molecular strategy, we have made significant progress in engineering drought tolerance in this important crop species.     

A Myosin XI Tail Domain Homologous to the Yeast Myosin Vacuole-Binding Domain Interacts with Plastids and Stromules in Nicotiana benthamiana

The actin cytoskeleton plays a role in mobility of many different organelles in plant cells, including chloroplasts, mitochondria, Golgi, and peroxisomes. While progress has been made in identifying the myosin motors involved in trafficking of various plant organelles, not all of the cargoes mobilized by different members of the myosin XI family have yet been identified. The involvement of myosins in chloroplast positioning and mitochondrial movement was demonstrated by expression of a virus-induced gene silencing （VIGS） construct in tobacco. When VIGS with two different conserved sequences from a myosin Xl motor was performed in plants with either GFP-labeled plastids or mitochondria, chloroplast positioning in the dark was abnormal, and mitochondrial movement ceased. Because these and prior obser- vations have implicated a role for myosins and the actin cytoskeleton in plastid and stromule movement, we searched for myosin tail domains that could associate with plastids and stromules. While a yellow fluorescent protein （YFP） fusion with the entire tail region of myosin XI-F was usually found only in the cytoplasm, we observed that an Arabidopsis or Nicotiana benthamiana YFP：：myosin XI-F tail domain homologous to the yeast myo2p vacuole-binding domain associated with plastids and stromules after transient expression in N. benthamiana. Taken together, these observations implicate myosin motor proteins in dynamics of plastids and stromules.     

低氧运动后不同恢复环境腓肠肌热休克蛋白70表达变化

目的：实验旨在观察急性间歇低氧（氧含量12．7％）跑台运动后不同恢复环境下大鼠腓肠肌热休克蛋白HSP70表达的时程变化。方法：雄性sD大鼠进行低氧环境下的急性间歇跑台运动,用Western blot方法检测低氧运动后即刻、低氧运动后低氧恢复及常氧氧恢复1d,2d,7d的大鼠腓肠肌HSP70蛋白表达水平。结果：急性间歇低氧运动后即刻HSP70蛋白表达水平开始升高。常氧恢复第1dHSP70蛋白表达水平显著高于常氧对照组,P〈0．05。第2d、第7d呈现先降低后升高的趋势;低氧恢复中HSP70蛋白表达水平均显著高于常氧对照组,P〈0．05。结论：急性间歇低氧运动后能诱导HSP70蛋白表达水平升高;低氧恢复过程中HSP70蛋白高水平表达维持的时间要比常氧恢复长。     

Protein-Repairing Methionine Sulfoxide Reductases in Photosynthetic Organisms： Gene Organization,Reduction Mechanisms,and Physiological Roles

Methionine oxidation to methionine sulfoxide （MetSO） is reversed by two types of methionine sulfoxide reducrases （MSRs）, A and B, specific to the S- and R-diastereomers of MetSO, respectively. MSR genes are found in most organisms from bacteria to human. In the current review, we first compare the organization of the MSR gene families in photosynthetic organisms from cyanobacteria to higher plants. The analysis reveals that MSRs constitute complex families in higher plants, bryophytes, and algae compared to cyanobacteria and all non-photosynthetic organisms. We also perform a classification, based on gene number and structure, position of redox-active cysteines and predicted sub-cellular localization. The various catalytic mechanisms and potential physiological electron donors involved in the regeneration of MSR activity are then de- scribed. Data available from higher plants reveal that MSRs fulfill an essential physiological function during environmental constraints through a role in protein repair and in protection against oxidative damage. Taking into consideration the ex- pression patterns of MSR genes in plants and the known roles of these genes in non-photosynthetic cells, other functions of MSRs are discussed during specific developmental stages and ageing in photosynthetic organisms.     

Plant Cell Wall Proteomics： Mass Spectrometry Data,a Trove for Research on Protein Structure/Function Relationships

Proteomics allows the large-scale study of protein expression either in whole organisms or in purified organelles. In particular, mass spectrometry （MS） analysis of gel-separated proteins produces data not only for protein identification, but for protein structure, location, and processing as well. An in-depth analysis was performed on MS data from etiolated hypocotyl cell wall proteomics ofArabidopsis thaliana. These analyses show that highly homologous members of multigene families can be differentiated. Two lectins presenting 93% amino acid identity were identified using peptide mass fingerprinting. Although the identification of structural proteins such as extensins or hydroxyproline/proline-rich proteins （H/PRPs） is arduous, different types of MS spectra were exploited to identify and characterize an H/PRP. Maturation events in a couple of cell wall proteins （CWPs） were analyzed using site mapping. N-glycosylation of CWPs as well as the hydroxylation or oxidation of amino acids were also explored, adding information to improve our understanding of CWP structure/function relationships. A bioinformatic tool was developed to locate by means of MS the N-terminus of mature secreted proteins and N-glycosylation.     

Alb4 of Arabidopsis Promotes Assembly and Stabilization of a Non Chlorophyll-Binding Photosynthetic Complex,the CF1CF0-ATP Synthase

All members of the YidC/Oxal/Alb3 protein family are evolutionarily conserved and appear to function in membrane protein integration and protein complex stabilization. Here, we report on a second thylakoidal isoform of Alb3, named Alb4. Analysis of Arabidopsis knockout mutant lines shows that AIb4 is required in assembly and/or stability of the CF1CF0-ATP synthase （ATPase）. alb4 mutant lines not only have reduced steady-state levels of ATPase subunits, but also their assembly into high-molecular-mass complexes is altered, leading to a reduction of ATP synthesis in the mutants. Moreover, we show that Alb4 but not AIb3 physically interacts with the subunits CF1β and CF0ll. Summarizing, the data indicate that AIb4 functions to stabilize or promote assembly of CF1 during its attachment to the membrane-embedded CF0 part.